The protective effects of humoral immunity are known to be mediated by a family of structurally related glycoproteins called antibodies. Antibodies initiate their biological activity by binding to antigens. Antibody binding to antigens is generally specific for one antigen and the binding is usually of high affinity. Antibodies are produced by B-lymphocytes. Blood contains many different antibodies, each derived from a clone of B-cells and each having a distinct structure and specificity for antigen. Antibodies are present on the surface of B-lymphocytes, in the plasma, in interstitial fluid of the tissues and in secretory fluids such as saliva and mucus on mucosal surfaces.
All antibodies are similar in their overall structure, accounting for certain similarities in physiochemical features such as charge and solubility. All antibodies have a common core structure of two identical light chains, each about 24 kilodaltons, and two identical heavy chains of about 55-70 kilodaltons each. One light chain is attached to each heavy chain, and the two heavy chains are attached to each other. Both the light and heavy chains contain a series of repeating homologous units, each of about 110 amino acid residues in length which fold independently in a common globular motif, called an immunoglobulin (Ig) domain. The region of an antibody molecule formed by the association of the two heavy chains is hydrophobic. Antibodies are known to cleave at the site where the light chain attaches to the heavy chain when they are subjected to adverse physical or chemical conditions. Because antibodies contain numerous cysteine residues, they have many cysteine—cysteine disulfide bonds. All Ig domains contain two layers of beta-pleated sheets with three or four strands of anti-parallel polypeptide chains.
Despite their overall similarity, antibody molecules can be divided into distinct classes and subclasses based on physiochemical characteristics such as size, charge and solubility, and on their behavior in binding to antigens. In humans, the classes of antibody molecules are: IgA, IgD, IgE, IgG and IgM. Members of each class are said to be of the same isotype. IgA and IgG isotypes are further subdivided into subtypes called IgA1, IgA2 and IgG1, IgG2, IgG3 and IgG4. The heavy chains of all antibody molecules in an isotype share extensive regions of amino acid sequence identity, but differ from antibodies belonging to other isotypes or subtypes. Heavy chains are designated by the letters of the Greek alphabet corresponding to the overall isotype of the antibody molecule, e.g., IgA contains α, IgD contains δ, IgE contains ε, IgG contains γ, and IgM contains μ heavy chains. IgG, IgE and IgD circulate as monomers. IgA molecules secreted through the epithelia into the mucosal lining of body cavities are homodimers whereas IgM molecules are pentamers. Circulating IgA exists mainly as a monomer. Multimeric forms of IgA and IgM are both stabilized by the so-called J chain. Secreted IgA (S-IgA) is produced by B cells residing in lamina propria and taken up by epithelial cells on the basolateral side through the poly-immunoglobulin receptor (plgR), transported through the epithelial cell and secreted into the mucosa on the luminal side. When the IgA:J chain:plgR complex is released, the plgR is cleaved by a protease and a part of the plgR molecule called the secretory component (SC) remains bound to the IgA:J chain complex. Thus, S-IgA is a complex consisting of IgA, the J chain, and the SC of which the two latter are covalently bound to the IgA molecule through disulphide bonds. S-IgA is very resistant to the proteolytic environment of the epithelial mucosa e.g. in the respiratory or the gastrointestinal tract, and as such make up the primary specific immune system in these sites. It has been demonstrated that S-IgA has an immunomodulating effect and may induce tolerance to the antigens they bind.
There are between 108 and 1010 structurally different antibody molecules in every individual, each with a unique amino acid sequence in their antigen combining sites. Sequence diversity in antibodies is predominantly found in three short stretches within the amino terminal domains of the heavy and light chains called variable (V) regions, to distinguish them from the more conserved constant (C) regions.
Immunoglobulin E (IgE) is responsible for so-called type 1 hypersensitivity which manifest itself as common diseases such as allergic rhinitis, allergic conjunctivitis, hay fever, allergic (extrinsic) asthma, bee venom allergy, and food allergy. Allergen-specific IgE is produced in excess in patients with IgE-mediated allergies. IgE circulate in the blood and bind to high-affinity Fc receptors for IgE on basophils and mast cells in blood, various tissues, or on mucosal surfaces. In most allergic responses, the allergens enter the body of a patient through inhalation, ingestion, or through the skin. The allergen molecules bind to preformed IgE already bound to the high-affinity receptor FcεRI on the surfaces of mast cells and basophils, resulting in the crosslinking of several IgE molecules and triggering the release of histamine and other inflammatory mediators causing the various allergic symptoms.
Among the tissues that are most susceptible to local IgE-mediated allergic reactions are the conjunctiva, the mucosa of the nasal cavity or the oropharynx (allergic rhinitis), the mucosal linings of the bronchial tract, and the gastrointestinal mucosa. Thus, allergens enter the respiratory tract through inhalation and get trapped on the mucosal surfaces of the nasal lining or the bronchial passages of the respiratory tract. Airborne allergens also get in contact with moist surfaces of eyes and ears and are retained on the mucosa. The mucosal tissues are densely populated with mast cells and allergens arriving at these sites may therefore bind IgE and activate mast cells.
The therapeutic principles and treatment modalities in the management of allergy have not changed substantially in recent years. Immunosuppressive drugs such as steroids for suppressing immune activities and bronchial dilators for relieving asthma symptoms have long been the main treatment modality for patients with allergic asthma. Desensitization immunotherapy is the most important novel therapy for severely affected patients, but the medical advances have been limited to refining the classification of the allergenic substances, improving diagnostic methods, and providing a better controlled and broader library of allergen extracts for immunotherapy. As for research, progress has been made in the identification and isolation of major allergenic components of allergenic substances. For example, the major allergic components of ragweed, house dust mites, and cat and dog dander and saliva have been identified. When the allergen particles, e.g. timothy grass pollen arrive to the airway mucosa they disintegrate into major and minor allergic components.
Antibodies have been suggested for a number of clinical treatments: Medimmune Inc. is studying the use of humanized anti-respiratory syncytial virus (RSV) monoclonal antibodies and markets a polyclonal anti-RSV human immunoglobulin product (RespiGam) isolated from human donor blood and used to treat RSV infection. Medimmune also markets CytoGam, an anti-CMV (cytomegalovirus) human immunoglobulin for the treatment of CMV infection. IDEC and Genentech are jointly performing clinical trials of a chimeric mouse-human monoclonal antibody (Rituximab) aimed at the CD20 antigen found on mature B cells and most non-Hodgkin's lymphoma tumors for the treatment of relapsed or refractory low-grade non-Hodgkin's lymphoma. GalaGen is studying the use of bovine polyclonal immunoglobulin (Diffistat-G) for treatment of Clostridium difficile antibiotic associated diarrhea. SmithKline Beecham and Schering-Plough are developing an anti-IL-5 monoclonal antibody which has been shown in clinical trials to prevent eosinophilic inflammation and airway constriction. An anti-IgE monoclonal antibody is being developed by Genentech to “switch-off” allergies. The antibody Rhu-Mab-E25, which is a humanized chimeric IgG, monoclonal antibody specific for a unique epitope on human high affinity IgE receptors (FcεRI), has been shown to reduce free IgE levels after the first administration by injection. It attenuated both early and late phase responses to inhaled allergens after multiple injections. Examples of antibodies used therapeutically also include a nebulized IgG (Sandoz), which is used intranasally against RSV; HNK20 (Oravax), an anti-RSV IgA; and 4B9 (Bristol Myers-Squibb), an anti-group B Streptococcus IgM monoclonal antibody Other therapeutically useful monoclonal antibodies include monoclonal anti-CD4 antibodies, anti-IL-2 antibodies and anti-IL-4 antibodies.
The immunotherapy of RSV infection using small particle aerosols of IgG has been disclosed by Piazza et al. (J. Infect. Dis., Vol. 166, pp. 1422-1424, 1992). In this study it was shown that a 15-minute exposure to an aerosolized 5% solution of IgG effected a 50-fold reduction in pulmonary virus. Brown (Aerosol Science and Technology, Vol. 24, pp. 45-56, 1996) discloses the use of antibodies as inhibitors or antagonists of cytokines to depress respiratory inflammatory diseases or allergen-induced asthmatic responses. Also mentioned is local respiratory delivery of pathogen-specific antibody for treatment of acute viral or bacterial respiratory infections.
Antibody liposomes, i.e., immunoliposomes, are disclosed by Maruyama et al. (Biochim. Biophys. Acta, Vol. 1234, pp. 74-80, 1995). Coating liposomes with antibody leads to enhanced uptake of the liposome by the reticuloendothelial system. Human monoclonal antibodies are known to be useful as anti-tumor agents. A mouse/human monoclonal IgG antibody specific for the Lewis Y antigen found on the surface of tumor cells is disclosed by Paborji et al. (Pharmaceutical Research, Vol. 11, No. 5, pp. 764-771, 1994). The use of antibodies in metered-dose propellant driven aerosols for passive antibody aerosol therapy against respiratory infections is suggested in Brown et al. (Journal of Immunological Methods, Vol. 176, pp. 203-212, 1994). Immune responses in the respiratory tract are of great importance for protection against infections of the respiratory system and for their involvement in respiratory allergies and asthma. Effective targeting of immunomodulating reagents including monoclonal antibodies to the respiratory tract is shown to be of benefit in increasing local immunity to respiratory pathogens or decreasing immune-mediated respiratory pathology. Inhaled immunoconjugates, immunoliposomes or immunomicrospheres have application in the lung as killers of cancer cells; (immunoconjugates) or, in the case of immunoliposomes and microspheres, as stealth delivery particles of a variety of therapeutic agents. An IgM anti-group B Streptococcus monoclonal antibody is disclosed by Gombotz et al. (Pharmaceutical Research, Vol. 11, pp. 624-632, 1994).
U.S. Pat. No. 5,670,626 proposes the use of monoclonal antibodies for the treatment of IgE-mediated allergic diseases such as allergic rhinitis, allergic asthma and allergic conjunctivitis by employing monoclonal antibodies to inhibit the entry of allergenic molecules into mucosal tissues. The binding of allergenic molecules by antibodies is assumed to inhibit the allergens from being taken up by mucosal epithelial cells
In certain clinical situations, the use of monoclonal antibodies is associated with specific disadvantages. Thus, monoclonal antibodies are directed against single antigenic epitopes. Therefore, if the target is of a complex nature presenting many different epitopes then the functional avidity of the monoclonal antibody may be low or lowered below a critical threshold allowing the target to escape elimination through immune recognition.
Also, because monoclonal antibodies are directed against single antigenic determinants, the density of the antibody targets on e.g. allergens may not be high enough to mediate elimination of the allergen. The efficient activation of complement similarly requires high target antibody densities which may not be achieved with single specificity monoclonal antibodies.
Thus, in the case of allergens, monoclonal antibodies are sub-optimal as they are directed against single epitopes. The majority of allergens are complex proteins, consisting of many protein and peptide epitopes, and existing in many variants. Thus, a single monoclonal antibody preparation cannot be expected to exhaustively cover more than a minority of the possible epitopes on an allergen, e.g. a pollen particle or proteins from cat dander. This means that if the desired clinical effect of an antibody can be characterized as a complete blocking of the available antibody epitopes, then a single monoclonal antibody will not be sufficient. Further, it an antibody preparation should preferably be developed against several homologous allergens from closely related allergens, e.g. pollens, or against several proteins from one allergen source e.g. animal dander, then a single monoclonal antibody will not meet the required efficacy.
Nevertheless, a paper by Schwarze and coworkers (Am. J. Resp. Crit. Care Med. Vol. 158, pp. 519-525, 1998) investigated the therapeutic efficacy of a monoclonal antibody directed against the major ragweed allergen Amb a I in a murine allergy model based on mice (Balb/c) sensitized and challenged with both Amb a I and whole ragweed extracts. It was demonstrated that administration of the monoclonal IgA antibody before allergen exposure decreased airway responsiveness to metacholine challenge, and decreased the number of pulmonary eosinophils and Amb a I-specific IgE levels in serum. Moreover, the study indicate that administration of IgA had an immunomodulatory effect implying that IgA treatment could have a long-term desensitizing effect on allergy. However, it must be stressed that this allergen model is based on the induction of allergy-like symptoms using a single allergen, Amb a I. Thus, the study does not take into account that the vast majority of allergies are caused by reactions towards a number of allergen proteins and epitopes derived from a single allergen particle, which emphasizes the need for a polyclonal antibody mixture in this regime of treatment. Furthermore, human allergy is profoundly more complex than the allergy-like symptoms induced in an inbred mouse strain (Inhal. Toxicol., Vol 12, pp. 829-62, 2000). Consequently, the potential usefulness of monoclonal antibodies as allergen blocking agents is limited. Finally, monoclonal antibodies may display cross-reactivity to antigenic structures of host cell tissue resulting in potential unwanted side effects. When this occurs the cross-reactivity cannot be removed by adsorption. Therefore a large number of different monoclonal antibodies may need to be produced in order to generate the desired combination of antigen specificity and target selectivity, and even so there still remains a significant risk of cross-reactivity towards endogenous self-antigens in a proportion of patients.
A separate issue is the generation of human anti-mouse antibody responses (HAMA). Conventional murine monoclonal antibodies are foreign proteins to the human recipient, and therefore a HAMA immune response is often elicited in the recipient, which may lead to unwanted side effects in addition to reduced treatment efficacy. In order to circumvent this problem, chimeric monoclonal antibodies possessing human constant (C) regions and murine variable (V) regions have been developed. Furthermore humanized monoclonal antibodies, where only the hypervariable complementarity determining region (CDR) is derived from mouse monoclonal antibodies and finally, so-called fully human monoclonal antibodies produced in mice transgenic for human immunoglobulin genes have been developed to avoid these problems. However, a potential for the generation of anti-idiotype antibody responses specific for the V-region specificity determining CDR still exists when injecting large amounts of monoclonal antibodies with identical V-regions.
For these reasons as outlined above, it may often be preferable to use polyclonal antibodies.
In WO 98/10776 it is theorized that phospholipase A2(PLA2) is involved in the pathogenesis of many diseases acting as an inflammatory mediator promoting chronic inflammation. Thus it is suggested to use serum reactive with at least one phospholipase A2 enzyme for the treatment of neoplasms in mammals. There is no suggestion to use polyclonal antibodies for blocking the uptake of an allergen by topical administration of an antibody binding to the allergen.
U.S. Pat. No. 4,740,371 describes a modification of allergen immunotherapy whereby an immune complex of the allergen and an antibody thereto is used for desensitization treatment, the antibody being present in molar excess with respect to the allergen to prevent an anaphylactic response. The purpose of the inclusion of the antibody in this treatment is to decrease the risk of allergic side effects such as anaphylactic shock to the desensitization treatment. The proportion of antibody to be added to the allergen is defined essentially by the neutralizing power of the antibody. Enough antibody must be used so that when the composition is administered, there is practically no allergic effect induced by the allergen. The adding of antibody to the allergen composition is solely a remedy to avoid side effects of the allergen exposure, the treatment still being an allergen immunotherapy.
There are several drawbacks of using conventional polyclonal antibodies in the treatment of allergy. First of all, polyclonal antibodies in the form of IgG purified from hyperimmune human serum is available in limited supply and in amounts insufficient for the treatment of allergic diseases and other common conditions. Also, gamma globulin preparations are expensive to produce, and display low efficacy due to their mixed nature containing an overwhelming majority of non-specific human serum immunoglobulin reactivities. Also, there exist a real risk of transmitting contaminating reagents, including infectious microorganisms (hepatitis virus, HIV, prions, others), or mitogens, cytokines and toxins. Finally, the variability between preparations remains a major problem. In order to solve the problem of supply, xenogeneic sources of polyclonal antibodies including serum from immunized non-human animals have been tested. However, such compositions may result in the generation of potent anti-xenoantibody responses, and carries a real risk of serious side effects such as anaphylactic shock or serum sickness, as well as the transmission of xenotropic infections.
U.S. Pat. No. 5,789,208 describes the use of a recombinant polyclonal antibody for vaccine therapy and prophylaxis to treat or prevent neoplastic diseases. The antibodies are used for boosting a patient's immune system for the possible later recognition of the antigen to which the antibody binds and thereby initiate an elimination reaction. The vaccination will have to be repeated to be effective. There is no suggestion to use polyclonal antibodies reacting with or binding to allergens in allergy treatment where the polyclonal antibodies should be administered completely differently before, during, or shortly after the patient has been exposed to an allergen.